Background/Purpose: BAFF inhibition is a new B cell targeted therapy approved for the treatment of moderately active SLE. Although BAFF regulates selection of naïve autoreactive B cells, belimumab only modest decreases autoantibody titers. To determine whether BAFF inhibition alters the quality of the autoantibody response by affecting selection of autoreactive B cells we generated NZW/BXSB.yaa (W/B) mice bearing the 3H9 site directed Ig heavy chain transgene. An extra copy of TLR7 conferred by the Yaa locus accelerates disease in the males. 3H9 associates with diverse light chains to generate anti-DNA and anti-cardiolipin specificity. We have previously shown that the autoreactive B cell response in 3H9.W/B mice uses predominantly Vk5-43 and Vk5-48 light chains that confer autoreactivity in their germline configuration and that such 3H9/Vk5-expressing B cells are rare in the naïve repertoire but are expanded in germinal centers (GCs).  Male W/B mice have defects in tolerance both at the GC entry checkpoint and in negative selection of B cells that acquire autoreactivity as a consequence of somatic mutation. We now wish to know whether BAFF inhibition influences either of these two checkpoints. 

Methods: To create a physiologic setting in which autoreactive B cells compete for survival with non-autoreactive B cells, we generated 50% 3H9/50% wt W/B bone marrow chimeras in which transferred 3H9. TLR7^+/+ or TLR7^+/-cells are GFP+ and can be easily identified.  To determine when negative selection of 3H9+ cells occurs, we enumerated the % 3H9+/GFP+ cells in immature bone marrow, transitional, marginal zone (MZ), follicular (Fo), GC and plasma B cells using flow cytometry. To investigate the effect of TLR7 dose and BAFF inhibitors on selection of the autoreactive B cell repertoire, we performed single cell PCR and sequencing to determine the repertoire of light chains associating with the 3H9 heavy chain in Fo, GC, and plasma cells. 

Results: When competition from wt B cells is provided, significant deletion of 3H9 B cells occurs at the immature and transitional stage in the bone marrow regardless of TLR7 status or BAFF inhibition. Relative enrichment of 3H9 B cells occurs in the GC TLR7^+/+ compared with TLR7^+/- chimeras. BAFF-R-Ig treatment significantly decreases B cell percentage and total numbers but does not decrease the percentage of immature or mature 3H9 B cells or mediate deletion of anergic cells. Data from single cell sorts of GFP+ B cells indicates comparable naïve B cell repertoires but a decrease in selection of high affinity anti-chromatin Vk5-48 /Jk4 light chains among TLR7^+/- and BAFF-R-Ig treated mice compared with TLR7^+/+ untreated controls.

Conclusion: Preliminary data indicates that when competition is provided, negative selection of naïve 3H9+ cells appears to be TLR7 and BAFF independent. In contrast, although autoreactive B cells still entered the germinal centers in mice treated with BAFF inhibition, modest changes were observed in germinal center selection. Future studies will explore the specific effects of BAFF inhibition on B cells that acquire autoreactivity in the GC as a consequence of somatic mutation.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/effects-of-baff-inhibition-on-b-cell-selection-in-murine-sle/